Lipid-conjugated polyamide compounds and related compositions and methods thereof

Abstract

In accordance with the present invention, there are provided lipid-conjugated polyamide compounds and related compositions and methods thereof. Lipid-conjugated polyamide compounds of the present invention are particularly useful as vehicles for delivering biologically active agents to a target site. In particular, the invention compounds are effective at facilitating the delivery of polynucleotides to cells. The present invention also provides a method for producing stable formulations of polynucleotides complexed with a delivery vehicle.

What is claimed:  
     
       1. A method of making a stable preparation of a polynucleic acid complexed with a delivery vehicle, said method comprising: 
       a) providing a polynucleic acid in a first liquid carrier as a dilute polynucleic acid solution that is substantially precipitant-free;  
       b) providing a delivery vehicle-forming compound in a second liquid carrier as a delivery vehicle solution that is substantially precipitant-free;  
       c) combining said dilute polynucleic acid solution with said delivery vehicle solution to form a dilute preparation of delivery vehicle/polynucleic acid complex; and  
       d) reducing the volume of said dilute preparation to form a stable preparation of delivery vehicle/polynucleic acid complex, wherein the concentration of polynucleic acid in said stable preparation is higher than the concentration of polynucleic acid in said dilute polynucleic acid solution, and wherein said stable preparation is substantially precipitant-free,  
       and wherein said delivery vehicle-forming compound is selected from (i) a lipid, selected from the group consisting of a phosphoglyceride, a glycosylglyceride, a sphingolipid, and a sterol, (ii) a polycationic non-lipid compound, and (iii) a lipid-conjugated polyamide compound comprising a repeating trimer of monomeric subunits and having the formula:                    
        where  
       m is an integer selected from 2 to about 48,  
       Ra is selected from the group consisting of —OH, —H, —SH, —COOH, sulfonyl, a C1-C8 aliphatic group or a C3-C12 aryl, arylalkyl, arylalkenyl, or arylalkynyl group, any of which may include at least one heteroatom selected from the group consisting of nitrogen, sulfur, oxygen, and phosphorus; and a lipid moiety, wherein said lipid moiety may be conjugated to a linker moiety;  
       each R 1  is independently selected from the group consisting of hydrogen, a C1-C8 aliphatic group or a C3-C12 aryl, arylalkyl, arylalkenyl, or arylalkynyl group, any of which may include at least one heteroatom selected from the group consisting of nitrogen, sulfur, oxygen, and phosphorus; and a lipid moiety, wherein said lipid moiety may be conjugated to a linker moiety, wherein at least one group R 1  is not hydrogen;  
       Rc is selected froth the group consisting of —OH, —H, —SH, —NH 2 , sulfonyl, hydrazine, a C1-C8 aliphatic group or a C3-C12 aryl, arylalkyl, arylalkenyl, or arylalkynyl group, any of which may include at least one heteroatom selected from the group consisting of nitrogen, sulfur, oxygen, and phosphorus; and a lipid moiety, wherein said lipid moiety may be conjugated to a linker moiety; and —each group W is selected from the group consisting of —CH 2 CH 2 —, CH 2 -phenyl-, CH 2 CH 2 O, —CH 2 CH═CH—, and —CR 2 R 3 —, where R 2  and R 3  are independently selected from the group consisting of hydrogen, a C1-C8 aliphatic group or a C3-C12 aryl, arylalkyl, arylalkenyl, or arylalkynyl group, any of which may include at least one heteroatom selected from the group consisting of nitrogen, sulfur, oxygen, and phosphorus; and a lipid moiety, wherein said lipid moiety may be conjugated to a linker moiety;  
       wherein any said aryl, arylalkyl, arylalkenyl, or arylalkynyl group having fewer than 5 carbon atoms further includes at least one heteroatom;  
       and wherein at least one of said groups Ra, Rc, R 1  for a single monomeric subunit, and W for a single monomeric subunit, comprises a lipid moiety.  
     
     
       2. The method according to  claim 1  wherein said delivery vehicle is a liposome. 
     
     
       3. The method of  claim 1 , wherein at least one group R 1  is a cationic sidechain selected from the group consisting of aminoalkyl, (S)α-methylethylenediamino, trimethylaminoethyl, guanidinoalkyl, aminobenzyl, pyridinium, cationic sidechains found on naturally occurring amino acids, and derivatives thereof. 
     
     
       4. The method of  claim 1 , wherein at least one group R 1  is a neutral sidechain selected from the group consisting of (S)-α-methylbenzyl, (R)-α-methylbenzyl, benzyl, phenethyl, naphthylmethyl, (S)-α-methylnaphthyl, (R)-α-methylnaphthyl, N-propylpyrrolidinone, cyclohexylmethyl, furfuryl, 3,4,5-trimethoxybenzyl, alkoxyalkyl, isoamyl, p-methoxyphenethyl, neutral sidechains found on naturally occurring amino acids, aid derivatives thereof. 
     
     
       5. The method of  claim 1 , wherein at least one group R 1  is an anionic side chain selected from the group consisting of carboxy, carboxy methyl, carboxy ethyl, benzoic acid, phosphates, sulfates, and derivatives thereof. 
     
     
       6. The method of  claim 1 , where each group W is —CR 2 R 3 —. 
     
     
       7. The method of  claim 6 , where R 2  and R 3  are hydrogen. 
     
     
       8. The method of  claim 7 , wherein R a  is selected from the group consisting of —OH, —H, —SH, —COOH, sulfonyl, and a lipid moiety, wherein said lipid moiety may be conjugated to a linker moiety. 
     
     
       9. The method of  claim 7 , wherein Rc is selected from the group consisting of —OH, —H, —SH, —NH 2 , sulfonyl, hydrazine, and a lipid moiety, wherein said lipid moiety may be conjugated to a linker moiety. 
     
     
       10. The method of  claim 7 , wherein Rc is a lipid moiety, which may be conjugated to a linker moiety. 
     
     
       11. The method of  claim 10 , wherein said lipid moiety is a sterol. 
     
     
       12. The method of  claim 7 , wherein Ra is a lipid moiety, which may be conjugated to a linker moiety. 
     
     
       13. The method of  claim 12 , wherein said lipid moiety is a phosphatidyl ethanolamine or a phosphatidyl propanolamine, having a phosphoglyceryl head group comprising two alkyl or alkenyl moieties having from 6 to about 25 carbon atoms. 
     
     
       14. The method of  claim 12 , wherein said lipid moiety is a sterol. 
     
     
       15. The method of claims  14 , wherein said sterol is a cholesterol. 
     
     
       16. The method of  claim 15 , wherein R 1   1  is a cationic sidechain, and R 1   2  and R 1   3  are neutral sidechains.